Although clinical responses in patients with metastatic melanoma occur after therapeutic vaccination, there is a need for improvement. Many potential target antigens have been identified, including shared tumor antigens and "class I" antigens that are unique to a given melanoma. However, more needs to be learned how best to deliver these antigens to induce a potent tumor-destructive immune response. Our approach is to first construct tumor vaccines that are therapeutically effective in mouse models and then to "translate" the findings to human melanoma and have developed several approaches that have efficacy in demanding mouse models. First, we showed that live or Mitomycin C-treated melanoma cells transfected to express anti-CD137 scFv can induce an immune response that can eliminate established tumors growing sc or in the lung. When compared against the same mouse tumors this approach is more effective than vaccination with tumor cells expressing CD137 ligand or administration of anti-CD137 MAb. Second, we demonstrated that transfection of tumor cells to express CD83 provides an alternative way to increase their immunogenicity for use as therapeutic vaccines. The mechanism is different from that engaged via CD137, and the two approaches can be combined for increased efficacy. Third, we have preliminary data indicating that melanoma cells transfected to express anti-CD40 scFv have efficacy as therapeutic vaccines in a model where the two other approaches did not work. Based on this information, we now propose experiments to learn more about mechanisms and to develop both tumor-cell based and gene-based vaccines that can cure mice with established melanomas of low immunogenicity. These findings will then be applied to construct human vaccines for in vitro evaluation towards the goal of identifying a vaccination strategy for Phase I trials in human patients with melanoma. Although all our planned studies will be performed with melanomas, the information gained is likely to be applicable also to other tumors. [unreadable] [unreadable]